Summary This is the second year of operation of this laboratory. The focus this year has been in onboarding and training of two new post-doctoral researchers, refinement of project trajectories, and generation of data for publications and presentations. Projects fall into two major categories: 1) Intrinsic electrical properties and synaptic inputs of medical olivocochlear (MOC) neurons in the brainstem, and 2) Influence of synaptic outputs of MOC neurons in the cochlea. Synaptic inputs of olivocochlear neurons in the brainstem and synaptic outputs in the cochlea Medial olivocochlear (MOC) neurons have cell bodies in the brainstem, where they receive synaptic inputs conveying sound information from the cochlea via the cochlear nucleus. The activation and inhibition of MOC neurons by this direct pathway from the ear is at early stages of investigation at the synaptic level. We aim to understand the diversity of synaptic inputs of MOC neurons, to fully understand how the neurons are activated, modulated, and how their properties may change to contribute to neuronal hyperactivity that has shown to occur in pathological situations such as in tinnitus or hyperacusis. Experimental accomplishments include development of a computational model of MOC neurons, including addition of intrinsic electrical conductances from experimental data collected from electrophysiological recordings of identified MOC neurons in our own laboratory. Specifically, these experiments investigate the role of the hyperpolarization-activated current Ih in affecting synaptic integration in MOC neurons. In a second project being primarily undertaken by a different lab member, we are also beginning a detailed investigation of synaptic inputs to MOC neurons in brainstem slices from mice. These experiments will elucidate the mechanisms of synaptic activation and inhibition of MOC neurons, which in turn drives their inhibition of mechanical activity in the cochlea, thus shaping the cochlear response to sound. Data is currently being generated that is expected to comprise two separate journal publications. Synaptic outputs of olivocochlear neurons in the cochlea MOC neurons project to the cochlea, where they decrease the mechanical movement of the basilar membrane by inhibiting cochlear OHCs. OHC activity enhances signaling to cochlear IHCs and shapes cochlear tuning curves and gain. MOC synapses onto OHCs are implicated in inhibiting OHC via coupling of cholinergic channel coupling to an SK potassium conductance. This effect is implicated in improved hearing in background noise, and protection of the cochlea against sound trauma. MOC neurons are also thought to release other neurotransmitters in the cochlea with poorly described effects on cochlear function. Equipment for this project has been installed and tested, and a new post-doctoral fellow is currently being trained in techniques of patch-clamp electrophysiology and imaging in dissected cochlear preparations. Behavioral assessment of olivocochlear function The medial olivocochlear system, a component of the final stage of the descending auditory system, is implicated in diverse effects on hearing including detection of salient sounds such as speech in a noise, protection of the cochlea against noise-induced trauma, and may have altered activity in pathological conditions such as tinnitus and hyperacusis. Recent reports from other laboratories provide contradictory evidence regarding the specific neurons that activate the MOC reflex. In a collaborative project with Dr. Rebecca Seal at the University of Pittsburgh, we have developed a mouse line that will provide more precise control of afferent signaling through type II spiral ganglion afferent neurons. We are in the planning stages of a collaborative project with Dr. Tracy Fitzgerald, Director of the NIDCD Animal Auditory Testing Core Facility, to test the function of the MOC system in control and in these mutant animals. These experiments will elucidate the pathways through which the MOC reflex is activated, distinguishing between inputs from type I vs type II spiral ganglion afferents. The experiments will employ measurements of distortion product otoacoustic emissions (DPOAEs), a test of OHC function. The change in DPOAE strength by contralateral suppression, a measure of MOC activity, will be used to assess the MOC function in control and mutant mice. These experiments will determine whether type II spiral ganglion afferent neurons indeed contribute to or modulate the MOC reflex. VGLUTs in OHCs, and the central innervation patterns of type II cochlear afferent neurons In a collaborative project with Dr. Rebecca Seal at the University of Pittsburgh, the specific vesicular glutamate transporters (VGLUTs) employed by OHCs to load glutamate into presynaptic vesicles for release onto spiral ganglion afferents was determined. This led to generation of mutant mice lacking the VGLUTs from either inner hair cells (IHC) or OHC, or both. Use of these mutant mice allows isolation of afferent signaling by either type of hair cell, in order to determine the unique contribution that each pathway makes to perception of sound. We used these different mouse lines in an experiment in which the animals were exposed to noise, then their cochlear nuclei assessed for activation of the immediate early gene c-Fos. We determined that the OHC-type II afferent pathway can indeed respond to acoustic signals, and evoke activation of neurons in the brainstem. A paper describing this work is currently in preparation.